The leaching of base cations in acidic soils can result in calcium (Ca2+) and magnesium (Mg2+) deficiencies, which are important for microbial cell function. We aimed to determine if microbial carbon use efficiency (CUE) and microbial biomass carbon (MBC) were limited in acidic soils due to a lack of base cations. Microbial CUE across a range of agricultural soils (n = 970; pHCa 3.4-7.9) treated with either deionised H2O (control) or a solution of 300 mM CaCl2 + 300 mM MgCl2 (+Base cations) was determined using a 14C radioisotope tracer approach. Our results showed that the addition of base cations significantly increased microbial CUE (by up to 20 %) at pHCa < 4.7; which coincided with a steep increase in exchangeable acidity. Base cation addition significantly increased MBC in nil-limed soils (pHCa 4.6) from 494 mg C kg-1 to 769 mg C kg-1 when plant residue was added, but not in limed soils (pHCa 6.2). Our findings indicate that the addition of base cations to highly acidic soils can increase microbial growth, thus aiding with carbon sequestration in these agricultural soils.